Alkali burn to the cornea is one of the most intractable injuries to the eye due to the opacity resulting from neovascularization (NV) and fibrosis. Numerous studies have focused on studying the effect of drugs on alkali-induced corneal injury in mouse, but fewer on the involvement of alkali-induced DNA methylation and the PI3K/AKT/mTOR signaling pathway in the mechanism of alkali-induced corneal injury. Thus, the aim of this study was to determine the involvement of DNA methyltransferase 3 B-madiated DNA methylation and PI3K/AKT/mTOR signaling modulation in the mechanism of alkali-induced corneal injury in a mouse model. To this end, we used bisulfite sequencing polymerase chain reaction and Western blot analysis, to study the effects of 5-aza-2′-deoxycytidine and 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one, which inhibit methyltransferase and PI3K respectively, on DNA methylation and expression of downstream effectors of PI3K related to corneal NV, including TSC1 and mTOR genes. The results showed that, after an intraperitoneal injection of rapamycin (2 mg/kg/day) for seven days, the alkali-induced opacity and NV were remarkably decreased mainly by suppressing the infiltration of immune cells into injured corneas, angiogenesis, VEGF expression and myofibroblasts differentiation; as well as by promoting corneal cell proliferation and PI3K/AKT/mTOR signaling. More significantly, these findings showed that epigenetic regulatory mechanisms by DNA methylation played a key role in corneal NV, including in corneal alkali burn-induced methylation modification and rapamycin-induced DNA demethylation which involved the regulation of the PI3K/AKT/mTOR signaling pathway at the protein level. The precise findings of morphological improvement and regulatory mechanisms are helpful to guide the use of rapamycin in the treatment of corneal angiogenesis induced by alkaline-burn.

由于新血管形成（NV）和纤维化导致的不透明性，碱烧伤角膜是对眼睛最棘手的伤害之一。许多研究集中于研究药物对小鼠碱诱导的角膜损伤的作用，但很少涉及碱诱导的DNA甲基化和PI3K / AKT / mTOR信号通路在碱诱导的角膜损伤的机制中的作用。因此，本研究的目的是确定在小鼠模型中碱诱导的角膜损伤的机制中涉及DNA甲基转移酶3 B酰化的DNA甲基化和PI3K / AKT / mTOR信号传导调节。为此，我们使用亚硫酸氢盐测序聚合酶链反应和蛋白质印迹分析，以研究5-氮杂-2'-脱氧胞苷和2-（4-吗啉基）-8-苯基-4H-1-苯并吡喃-4-一，TSC1和mTOR基因。结果表明，腹膜内注射雷帕霉素（2 mg / kg /天）7天后，碱诱导的不透明性和NV显着降低，主要是通过抑制免疫细胞向受损角膜的浸润，血管生成，VEGF表达和肌成纤维细胞分化；以及通过促进角膜细胞增殖和PI3K / AKT / mTOR信号传导。更重要的是，这些发现表明，DNA甲基化的表观遗传调控机制在角膜NV中起关键作用，包括在角膜碱烧伤诱导的甲基化修饰和雷帕霉素诱导的DNA去甲基化中，涉及PI3K / AKT / mTOR信号通路的调控。蛋白质水平。